Spinal fusion methods and devices

ABSTRACT

Methods, devices and compositions for fusing adjacent vertebrae, and otherwise localizing bone growth, are provided. In one form of the invention, a method for fusing adjacent vertebrae includes preparing a disc space for receipt of an intervertebral disc implant in an interwertebral disc space between adjacent vertebrae, inserting the implant into the intervertebral disc space and providing an osteoinductive composition that includes an osteoinductive factor in a pharmaceutically acceptable carrier. The carrier is advantageously substantially impermeable to efflux of the osteoinductive factor and is released as the carrier is resorbed or biodegraded. Preferred carriers include a hardened, resorbable carrier, such as a calcium phosphate cement that retains at least about 50% of the osteoinductive factors greater than about 2 days. Preferred osteoinductive factors are growth factors and include bone morphogenctic proteins and LIM mineralization proteins. In alternative forms of the invention, the method may be performed without utilization of a load-bearing spinal implant by disposing the osteoinductive composition in the disc space. The method is advantageously performed on lumbar vertebrae by a posterior approach. Intervertebral fusion devices and methods for their preparation are also provided.

CROSS-REFERENCE TO RELATED APPLICATION

The present application claims the benefit of U.S. ProvisionalApplication Ser. No. 60/424,794, filed on, Oct. 24, 2000, which ishereby incorporated by reference in its entirety.

BACKGROUND OF THE INVENTION

The present invention relates generally to methods and devices forstabilizing the spine. More specifically, the invention provides methodsand devices for fusing adjacent vertebrae and for localizing spinal bonegrowth.

Back pain affects millions of individuals and is a common cause ofdisability for the middle-aged working population. A frequent cause ofback pain is rupture or degeneration of intervertebral discs.

Intervertebral discs, located between the endplates of adjacentvertebrae, stabilize the spine, distribute forces between vertebrae andcushion vertebral bodies. An intervertebral disc includes the nucleuspulposus, a gelatinous component that is surrounded and confined by anouter, fibrous ring, called the annulus fibrosus. In a healthy,undamaged spine, the annulus fibrosus prevents the nucleus pulposus fromprotruding outside the disc space.

Spinal discs may be displaced or damaged due to trauma; disease, oraging. Disruption of the annulus fibrosus allows the nucleus pulposus toprotrude into the vertebral canal; a condition commonly referred to as aherniated or ruptured disc. The extruded nucleus pulposus may press on aspinal nerve, which may result in nerve damage, pain, numbness, muscleweakness and paralysis. Intervertebral discs may also deteriorate due tothe normal aging process or disease. As a disc dehydrates and hardens,the disc space height will be reduced leading to instability of thespine, decreased mobility and pain.

In many instances, the only relief from the symptoms of these conditionsis a discectomy, or surgical removal of all or a portion of anintervertebral disc followed by fusion of the adjacent vertebrae. Theremoval of the damaged or unhealthy disc will allow the disc space tocollapse. Collapse of the disc space can cause instability of the spine,abnormal joint mechanics, premature development of arthritis or nervedamage, in addition to severe pain. Pain relief afforded by a discectomyand arthrodesis requires preservation of the disc space and eventualfusion of the affected motion segments.

One solution to the stabilization of an excised disc space is to fusethe vertebrae between their respective endplates. Typically anosteoinductive material is implanted at the treatment site to promotespinal fusion. Success of the discectomy and fusion procedure requiresdevelopment of a contiguous growth of bone to create a solid masscapable of withstanding the compressive loads on the spine for the lifeof the patient.

Additionally, several metal spacers have been developed for implantationinto a disc space and can be used to promote fusion. Medtronic SofamorDanek, Inc., (Memphis, Tenn.) markets a number of hollow spinal cages,and a wide variety of other such cages are known in the art. Forexample, U.S. Pat. Nos. 5,015,247 and 5,984,967 to Michelson et al. andZdeblick et al., respectively, disclose threaded spinal cages. The cagesare hollow and-can be filled with osteoinductive material, such asautograft, allograft and/or material isolated from the grafts. Aperturesdefined in the cages communicate with the hollow interior to provide apath for tissue growth between the vertebral endplates.

Such implants have been positioned in vivo by medical procedures wellknown in the art, including anterior and posterior approaches. Incertain instances, it is possible that the osteoinductive material thatincludes an osteoinductive factor may diffuse, or otherwise migrate,from the implant into undesired locations, which may result in boneformation in these locations. For example, the osteoinductive materialmay diff-use out of the cage, or other implant, and may form bone insidean adjacent hematoma, or tissue, such as fibrous scar tissue. The can bean increased risk of hematoma formation with posterior lumbar interbodyfusion (PLIF) or transforamiinal lumbar interbody fusion procedure,because the blood released during these procedures can pool in thespinal canal or foramen space. Scar tissue formation from pooling bloodfrom prior surgeries is also more prone in revision PLIF or TLIFprocedures. There is therefore a need for methods for fusing adjacentvertebrae and osteoinductive compositions that aid in reducing formationof bone tissue in unwanted, or otherwise undesired, locations.

In light of the above described problems, there is a continuing need foradvancements in the relevant field, including improved methods fortreating orthopedic injuries and defects, osteogenic compositions anddevices relating to enhancing spinal fusion. The present invention issuch an advancement and provides a wide variety of benefits andadvantages.

SUMMARY OF TH INVENTION

It has been discovered that blending osteoinductive compositions with aslow release carrier can effectively reduce bone formation in undesiredlocations during spinal fusion procedures. For example the slow releasecarrier can inhibit migration of the entrained osteoinductivecomposition to tissue adjacent the treatment site, for example, sites ofhematoma, scar tissues, or other fibrous tissues that are a distancefrom, or adjacent to, the desired site for fusion. Accordingly, oneaspect of the invention provides methods for fusing adjacent vertebrae,and otherwise localizing bone growth, in an interbody fusion procedure.The method is particularly advantageous for treatment sites that alreadyexhibit a localized hematoma or scar tissues or exhibit a clinicalpredisposition for such. During the disc space preparation, a hematomasite or scar tissue site can be exposed or evaluated for apredisposition for bone tissue growth induced by a diffusibleosteoinductive factor. In one embodiment, an osteoinductive compositionand a carrier composite can be formulated to promote and limit bonegrowth to a desired treatment site. In another embodiment, an implantcan be advantageously formulated and configured to retain anosteoinductive composition. The osteoinductive composition is thereforeprovided to and carried by the implant minimizing undesirable migrationof the osteoinductive composition from the implant.

In other forms the invention provides minimally invasive methods forfusing adjacent vertebrae. The method comprises preparing a disc spacefor receipt of an osteoinductive composition and the osteoinductivecomposition is inserted into the prepared disc space, withoututilization of a load-bearing spinal implant. Such methods may be usedin conjunction with instrumentation of the spine, such as anterior orposterior instrumentation with rods, plates and the like.

A composition for use in the invention includes a carrier and aneffective amount of an osteoinductive material or an osteoinductivefactor. The osteoinductive material can be entrapped or entrained withinthe carrier. The carrier is preferably substantially impermeable toefflux of the osteoinductive factor. In one embodiment, the osteogenicmaterial is released from the carrier as the carrier is degraded orresorbed. In one embodiment, bone formation can be substantiallyconfined to the original volume or space occupied by the carrier,osteogenic material. Migration of the osteogenic material to thehematoma site or scar tissue site is significantly reduced oreliminated. In another embodiment of the invention, the carrier is aresorbable cement and the osteogenic material is an osteoinductivefactor or bone morphogenetic protein.

In other forms the invention provides methods for performing a posteriorlumbar interbody fusion or a transforaminal lumbar interbody fusion,wherein the selected disc for treatment is a lumbar disc.

Intervertebral fusion devices that include a spinal implant and theosteoinductive compositions described above are also provided, as aremethods of preparing the fusion device.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a series of scanned images of CT scans of three axial slicesthrough a Rhesus monkeys' vertebra that were treated by posterolateraltransverse process fusion using rhBMP-2 in a standard Etex calciumphosphate cement carrier. The CT scans were taken at 2, 4 and 6 monthsafter implantation.

FIG. 2 is a series of scanned images of CT scans of the three axialslices through a Rhesus monkeys' vertebra that were treated byposterolateral transverse process fusion using rhBMP-2 in a modifiedEtex calcium phosphate cement carrier. The CT scans were taken at 2, 4and 6 months after implantation.

FIG. 3 is a series of scanned images of the whole spine X-ray of amonkey treated as in FIG. 1.

FIG. 4 is a series of scanned images of the whole spine X-ray of amonkey treated as in FIG. 2.

FIG. 5 is a graph illustrating rhBMP-2 release kinetics from Etexcement.

FIG. 6 is a graph illustrating the retention profile of rhBMP-2 in Etexcement.

DESCRIPTION OF THE EMBODIMENTS

For the purposes of promoting an understanding of the principles of theinvention, reference will now be made to embodiments and specificlanguage will be used to describe the same. It will nevertheless beunderstood that no limitation of the scope of the invention is therebyintended, such alterations and further modifications of the invention,and such further applications of the principles of the invention asillustrated herein, being contemplated as would normally occur to oneskilled in the art to which the invention relates.

The present invention relates to methods and devices for treatingadjacent vertebrae. In preferred forms of the invention, methods areprovided for fusing adjacent vertebrae by a posterior or transforaminalinterbody fusion approach, such as a posterior lumbar interbody fusion(PLIF) or transforaminal lumbar interbody fusion CMIF) approach. Methodsfor localizing bone formation to a pre-selected location in anintervertebral disc fusion procedure are also provided. In certain formsof the invention, the methods described herein may also be performedposterolaterally or anterolaterally. In preparing the disc space in aposterior or transforaminal interbody fusion procedure, tissue can beexposed that is susceptible to or is predisposed to undesirable bonetissue growth. Desirable bone tissue growth is promoted by blending,dispersing or otherwise entraining a selected osteoinductive materialwith a carrier that does not allow substantial efflux of the material oran active portion of the material from the carrier. In one form, theosteoinductive material slowly diffuses out of the carrier. In anotherform the carrier does not release a substantial portion of theosteoinductive material, but rather as the carrier degrades and/or isbioabsorbed and gradually exposes the entrained osteoinductive materialto the treatment site.

Tissue that has a predisposition to bone growth includes scar tissuethat can preexist from a previous surgery or other incident disease(s)or injury causing formation thereof. The scar tissue can form, or beotherwise located, at a site a distance from, typically adjacent to, adesired site of bone formation. For example, scar tissue may have formedbecause of fibroblast invasion into a blood clot or hematoma from aprior surgery.

Furthermore, a hematoma can be formed during or a result of the currentsurgery described herein or a prior surgical procedure, disease and/ortrauma The hematoma can be created, or otherwise located a distancefrom, or adjacent to, the desired bone formation site. For example,tissue injured or bruised during the surgical procedure seeps blood andother fluid, and this tissue can continue to seep blood/fluid subsequentto surgical closure. This blood/fluid can pool and collect in tissuecavities and pockets such as in the spinal canal and foramen space.

It is often desirable to minimize diffusion of osteoinductivecompositions into a hematoma or scar tissue because these compositionscan induce calcification of the hematoma or scar tissue into bonetissue.

In one aspect of the invention, methods for fusing adjacent vertebrae,and otherwise localizing bone growth, are provided. In one form of theinvention, the vertebrae are fused by a PLIF or TLIF procedure. Apatient is first prepared for the surgical procedure. For example, thepatient is properly positioned on the operating table, typically in aprone position with their pelvis parallel to the floor. Access to thevertebral level to be fused, such as the selected lumbar disc, is thengained utilizing surgical methodology and tools. The intervertebral discspace and adjacent vertebrae are then prepared for receipt of anintervertebral disc implant or spacer. The spacer is prepared orconfigured to retain an advantageous osteoinductive composition. Incertain forms of the invention, a spacer is not utilized and theosteoinductive composition is injected, or otherwise disposed, into theintervertebral disc space, thus eliminating the open surgical proceduresoften necessary for spinal implant placement.

The present invention can combine minimally invasive surgery methods.The disc space may be prepared by minimally-invasive methods known tothe skilled artisan, typically by making a small incision in thepatient, such as no larger than about 30 mm, and inserting a cannulapercutaneously into the patient through which the necessary tools can bedelivered to, and manipulated at, the surgical site. The osteoinductivecomposition can be injected into the disc space.

Moreover, as desired or deemed medically prudent, instrumentation of thespine, including rods and plates, can advantageously be utilized incertain forms of the invention to maintain or restore desired disc spaceheight and prevent disc space collapse after surgery during the fusionprocess.

The osteoinductive composition is combined with a pharmaceuticallyacceptable slow-release carrier. A preferred carrier is selected thatallows a slow release of the osteoinductive factor. “Slow release” isdefined herein to mean release of the osteoinductive factor at a ratethat substantially reduces release of the osteoinductive factor from thecarrier and thus substantially reduces migration, or diffusion, of theosteoinductive factor to tissue a distance from, typically adjacent to,the carrier or implant. The distant site can include scar tissue orother fibrous tissue, a hematoma, or other collection of blood cells ortissue, which can exhibit a propensity for bone tissue growth. Thus,“slow release” as defined herein also means release of theosteoinductive composition at a rate that substantially decreases bonetissue formation in undesired locations. Slow release also includes arate of release wherein the half-life for release of the osteoinductivecomposition from the carrier is typically greater than about 2 days,preferably at least about 4 days, more preferably at least about 7 daysand still more preferably at least about 14 days. With respect to theosteoinductive composition, the half-life refers to the amount of timeit takes 50% of the mass of composition to be released from the carrier.In a one embodiment, the osteoinductive composition(s) are completedreleased from the carrier within about 8 to about 12 weeks.

In other forms, the carriers of the present invention are biodegradableand exhibit a half-life for maintaining their integrity. The carrier'shalf-life is the time period in which one half of the carrier's mass hasbeen degraded or absorbed. In a one embodiment, the carrier's half-lifeis typically greater than about 2 days post implantation, preferably atleast about 4 days, more preferably at least about 10 days and stillmore preferably at least about 14 days. Further, in other embodiments,the carrier is selected or formulated such that it is not completelydegraded or its mass or volume approximates zero until at least about 8weeks post implantation; more preferably the selected carrier is notcompletely degraded or its mass/volume reduced to zero before about 16weeks post implantation.

The carrier can be selected and/or formulated to be flowable orinjectable at a high temperature and which hardens at a lowertemperature. The high temperature should not be at such a high level tocause tissue damage, and therefor, the high temperature is typicallyselected to be below about 60° C., more preferably below about 50° C.,and still more preferably below about 45° C. The low temperature shouldbe sufficiently high so the carrier maintains its selected conformationat body temperature and can take into account higher than normal bodytemperature levels caused by fever from infections or otherphysiological phenomenon. The low temperature level can be selected tobe at least about 37° C. more preferably at least about 40° C.

In alternative embodiments, polymer based carriers are selected. Thepolymer based carriers are preferable a polymer matrix having pores suchas can be found in sponge-like matrixes. The polymeric material can be ashape memory polymeric material as described in U.S. patent applicationSer. No. 09/696,389, 09/696,146 or 09/696,715 all filed on Oct. 25,2000. The polymeric material can be modified to slowly release anosteoinductive composition. For example, increasing the crosslinkingbetween polymeric chains, combining the polymer with a collagen, gelatinor carboxymethylcellulose, or glycan, can serve to entrap anosteoinductive composition within the polymer matrix.

A wide variety of carriers may be used in the invention. Suitablecarriers include polymers, such as, polylactic acid, polyglycolic acid,alternating copolymers of polylactic acid and polyglycolic acid,polyethylene glycol. These polymers may be formed into a matrix such asa sponge with voids for the infiltration with the osteoinductivematerial. Optionally, these polymers can be combined with one or more ofcarboxymethylcellulose hyaluronic acid, glycans such asglycosaminoglycans, gelatin, and/or collagen to effect suitable releaseprofiles. The release rates from these polymers can be furtherinfluenced by chemical modification such as inducing and/or increasingpolymer chain length and/or cross-linking, forming semi-interpenetratingpolymer networks (SIPN) or interpenetrating polymer networks (IPN), starpolymers polymer complexes and blends or polymer alloys and acombination thereof. Further, carboxymethylcellulose hyaluronic acid,glycans such as glycosaminoglycans, gelatin, and/or collagen can bemodified to exhibit suitable release profiles by increasing the densityof these compositions. Other carriers suitable for use with thisinvention include resorbable cements such as calcium phosphate,tricalcium phosphate and hydroxy apatite based cements. The resorbablecements may be substantially amorphous materials having the slow releaseproperties described herein. In one form the carriers are formulated tohave a higher affinity for selected osteoinductive compositions. Thesecarriers can then be combined and/or compounded with the selectedosteogenic compositions.

In one form, the carrier is provided as a calcium phosphate cement. Suchcalcium phosphate cements are preferably synthetic calcium phosphatematerials that include a poorly or low crystalline calcium phosphate,such as a low or poorly crystalline apatite, including hydroxyapatite,available from Etex Corporation and as described, for example, in U.S.Pat. Nos. 5,783,217; 5,676,976; 5,683,461; and 5,650,176, and PCTInternational Publication Nos. WO 98/16268, WO 96/39202 and WO 98/16209,all to Lee et al. By use of the term “poorly or low crystalline” ismeant to include a material that is amorphous, having little or no longrange order and/or a material that is nanocrystalline, exhibitingcrystalline domains on the order of nanometers or Angstroms. Thecalcium: phosphate ratio of the carrier is typically in the range ofabout 0.3 to about 0.7, more preferably about 0.4 to about 0.6.

In another form the carrier can be modified to exhibit a substantiallyclosed porous structure. The osteoinductive material is preferablecombined or blended with the carrier material or precursor prior tomodification carrier into a substantially closed matrix. Theosteoinductive material becomes trapped within the inner cells of thematrix. In use the carrier material slowly erodes, and as the carriermaterial erodes, the inner cells entraining the osteoinductive materialare exposed. The exposed cells release the osteoinductive materialcontained therein. The biodegradation rate of the carrier can be variedas desired to vary the release rate of the osteoinductive material.

Utilizing the carrier described herein, bone formation is advantageouslyconfined to the volume of the carrier. The bone that forms may thus beconfigured or otherwise shaped as the original shape of the carrier uponimplantation. A carrier may confine or otherwise entrap theosteoinductive factor within the carrier so that the factor will besubstantially released as the carrier is resorbed. Stated alternatively,the carrier will advantageously be substantially impermeable to effluxof the osteoinductive factor. It is further preferred that the carrieris selected so that substantially no osteoinductive factor migrates, orotherwise diffuses, into areas of unwanted bone formation as describedabove, or the amount of osteoinductive factor that may migrate into suchareas will not be sufficient to substantially induce bone tissuegeneration.

A wide variety of osteoinductive factors may be used in the osteogeniccomposition, including bone morphogenetic proteins (BMPs), LIMmineralization proteins (LMPs), including LMP-1, growth differentiationfactors (GDF), cartilage-derived morphogenic proteins (CDMP) and othergrowth factors such as epidermal growth factors, platelet-derived growthfactors, insulin-like growth factors, fibroblast growth factors andtransforming growth factors, including TGF-β and TGF-α, and combinationsthereof A wide variety of bone morphogenetic proteins are contemplated,including bone morphogenetic proteins designated as BMP-2 throughBMP-18, heterodimers thereof and combinations thereof. Proteins can berecombinant proteins, such as, recombinant human proteins. Suitablerecombinant human bone morphogenetic proteins (rhBMPs) include rhBMP-2and rhBMP-7.

BMPs are available from Genetics Institute, Inc., Cambridge,Massachusetts and may also be prepared by one skilled in the art asdescribed in U.S. Pat. No. 5,187,076 to Wozney et al.; U.S. Pat. No.5,366,875 to Wozney et al.; U.S. Pat.No. 4,877,864 to Wang et al.; U.S.Pat. No. 5,108,922 to Wang et al.; U.S. Pat. No. 5,116,738 to Wang etal.; U.S. Pat. No. 5,013,649 to Wang et al.; U.S. Pat. No. 5,106,748 toWozney et al.; and PCT Patent Nos. WO93/00432 to Wozney et al.;WO94/26893 to Celeste et al.; and WO94/26892 to Celeste et al. All bonemorphogenic proteins are contemplated whether obtained as above orisolated from bone. Methods for isolating bone morphogenetic proteinfrom bone are described, for example, in U.S. Pat. No. 4,294,753 toUrist and Urist et al., 81 PNAS 371, 1984.

The osteoinductive composition may include the osteoinductive factors,or nucleotide sequences that encode the respective osteoinductivefactors, so that the osteoinductive factor may be produced in vivo, in apharmaceutically acceptable carrier. The nucleotide sequences can beoperably linked to a promoter sequence and can be inserted in a vector,including a plasmid vector. A nucleic acid sequence can be “operablylinked” to another nucleic acid sequence when it is placed in a specificfunctional relationship with the other nucleic acid sequence.

In other embodiments, cells may be transformed with nucleotide sequencesencoding the osteoinductive factor and the osteoinductive compositionwill then include the transformed cells in a pharmaceutically acceptablecarrier. In other forms, the osteoinductive composition includes a virussuch as, for example, an adenovirus capable of eliciting intracellularproduction of a LIM mineralization protein.

When utilizing a resorbable cement carrier, a cell-sustaining componentis further included in the carrier. The cell-sustaining component is onethat provides nutrients to the cells so that they are able to producethe osteoinductive factor. The cell-sustaining component is alsoselected so that it does not substantially alter or otherwise modify therate at which the carrier is resorbed or the rate at which theosteoinductive factor is released. Such cell-sustaining componentsinclude collagen, and various cell culture media utilized for ex vivocell culture, including an infusible media such as normal salinesupplemented with about 5% human serum albumen (HSA), Dulbecco'sModified Eagle's medium (DMEM), or RPMI 1640 supplemented with fetalbovine serum or serum-free medium formulations such as the X VrVOproducts, or the components include a combination thereof. In yet otherforms of the invention, the nucleotide sequences may be combineddirectly with the carrier for delivery.

The amount of osteoinductive factor included in the carrier, and theamount applied to the treatment site, is typically an amount effectivein forming new bone and eventual fusion of adjacent vertebrae. Thisamount will depend on a variety of factors including the nature of theosteoinductive factor, the osteoinductive potential of the factor, andthe nature of the carrier, but will typically be about 0.5 mg BMPTmlcarrier to about 4 mg BMP/ml carrier (corresponding to a weight ratio ofBMP:dry carrier of about 1:2000 to about 1:250). The compositions mayinclude about 1 mg BMP/ml carrier to about 3 mg BMP/ml carrier(corresponding to a weight ratio of BMP:dry carrier of about 1:1000 toabout 1:333), but typically include at least about 2 mg BMP/ml carrier(corresponding to a weight ratio of BMP: dry carrier of at least about1:500). The amount of the osteoinductive composition applied to thefusion site will also vary, but will typically be sufficient to deliverabout 2 mg BMP to about 40 mg BMP, preferably about 4 mg BMP to about 20mg BMP, and typically at least about 12 mg BMP.

Additionally, a wide variety of different implants or spacers can beused with the present invention. The implants or spacers may beconfigured to retain an osteoinductive composition as described herein.Implants may include chambers, channels, pores or other spaces in whichthe osteoinductive composition may be packed, placed or otherwiseretained. The implants or spacers can be either resorbable/biodegradableor nonresorbable/biodegradable. Further the implants can beeintervertebral fusion devices, such as cages having a chamber therein,and optionally with end caps to further aid in retaining theosteoinductive composition. Examples of suitable implants may be foundin U.S. Pat. Nos. 4,961,740; 5,015,247; 5,423,817, PCT Applications No.PCT/US01/08193 and PCT/US01/08073, and published PCT Application WO99/29271.

Intervertebral fusion devices including the spinal implant andosteoinductive compositions described above are also provided which mayadvantageously be used to localize bone to desired areas as describedherein during an intervertebral fusion procedure, especially a PLIF or aTLIF procedure.

In methods described herein utilizing a spinal implant, theosteoinductive composition is preferably disposed in, on or is otherwiseassociated with, the spinal implant described herein prior to insertingthe implant in the intervertebral disc space. The ossteoinductivecomposition can be disposed in one or more chambers in an implant priorto inserting the implant into the intervertebral disc space. It isfurther realized that the osteoinductive composition may be retained orotherwise disposed on or in the implant while positioning the implant inthe disc space or after it is so positioned. When disposing thecomposition on or in the implant after it is disposed in the disc space,it is preferred to utilize a carrier in a flowable form, which willpreferably harden at about body temperature, although it is realizedthat the carrier may be in a wide variety of forms prior to disposingthe carrier on or in the implant, including a flowable or non-flowable,hardenable or hardened form, as long as it will ultimately be in ahardened form or state in vivo at pharmacological temperature, pH, andin selected body fluids such as is found proximal to bone tissue andconnective tissue.

Reference will now be made to specific examples illustrating thecompositions, methods and devices described above. It is to beunderstood that the examples are provided to illustrate preferredembodiments and that no limitation to the scope of the invention isintended thereby.

EXAMPLE 1 Single-level Posterolateral Fusion in Rhesus Monkeys

This example shows that posterolateral fusions performed in Rhesusmonkeys with the bone substitute compositions described herein resultedin new bone formation that was confined to the volume occupied by thebone substitute compositions.

Animals and Experimental Design

Posterolateral transverse process fusions in 2 groups of 2 Rhesusmonkeys were performed. One of the groups received rhBMP-2 in a carrierof standard α-bone substitute material (standard α-BSM®), a commerciallyavailable calcium phosphate cement purchased from Etex Corp., Cambridge,Mass. This standard material has a microporosity of 40%. The other groupreceived rhBMP-2 in a carrier of modified α-BSM® (a bone substitutematerial from Etex with a porosity greater than 40%). In this secondgroup, the left side of the spine was treated with the rhBMP-2 in acarrier of Δ-BSM® having 80% porosity (the porosity was increased byincreasing the liquid content) and the right side of the spine wastreated with α-BSM® having a porosity of about 80% from addition ofcollagen fibers to the standard α-BSN®. The extent of fusion wasobserved by Computer Tomography (CT) and X-ray analyses.

Preparation of BMP/Carrier

The rhBMP-2 was supplied in a buffer solution, pH 4.5 from GeneticsInstitute, Cambridge Mass. The rhBMP-2 solution was withdrawn from theprovided vial with a needle and syringe and injected into a plasticmixing “bulb” containing the α-BSM® dry powder. The powder was thenmixed by hand by kneading the plastic bulb for about 2-3 minutes until amixture having a putty-like consistency is obtained. The tip of the bulbwas cut off and the putty material applied, or otherwise administered,to the respective spinal posterolateral fusion site. The rhBMP-2concentration was 2.1 mg BMP/ml of carrier. A composite comprising about15 ng of BMP and about 7 ml of the putty-like material was used on eachside of the spine.

Results

It was found that fusion was achieved in all animals studied and thatnew bone formation was confined to the shape of the bone substitutematerial implanted across the transverse process. FIG. 1 is a series ofscanned images of CT scans of taken at three different levels throughthe treated transverse process site at 2, 4 and 6 months postimplantation for the first set of monkeys treated with rhBMP-2 in astandard Etex carrier. Similarly, FIG. 2 is a series of scanned imagesof CT scans of taken at three different levels through the treatedtransverse process site at 2, 4 and 6 months post implantation for thesecond set of monkeys treated with rhBMP-2 in a modified Etex carrier.It can be seen after analyzing FIGS. 1 and 2, that the shape and size ofthe fusion mass remains the same over time, indicating that the carrierretains the BMP within its matrix. As the carrier resorbs from theoutside surface inward, it is replaced by new bone, thus resulting inprecisely controlled bone formation.

FIGS. 3 and 4 are scanned images of X-rays taken at 1, 2, 4.5 and 6months post implantation of the spinal column of the monkeyscorresponding to the CT scans in FIGS. 1 and 2, respectively. As seen inFIGS. 3 and 4, the α-BSM® resorbs over time and is replaced by new boneacross the transverse process.

EXAMPLE 2 Pharmacokinetic Study of the Release of rhBMP-2 From α-BSM andACS

The release kinetics for rHBMP-2 from α-BSM and ACS evaluated in arabbit ulna osteotomy. A 125I-rhBMP-2/α-BSM or 125IrhBMP-2/ACS productwas surgically implanted in a rabbit ulna osteotomy. Assessment of theradioactivity at the implant site were made as soon as possiblefollowing surgery. Additionally assessments were made periodicallythereafter including at 1, 2, 3, 4, 7, 14 and 21 days after surgery.

The rhBMP-2 was radiolabeled with ¹²⁵I using the iodogen technique. Thefollowing a typical procedure. An 80 μg/mL solution of iodogen reagent(Pierce, Rockford, Ill.) was prepared in chloroform. An aliquot of thissolution (50 μL) was placed into a micro-eppendorf tube and evaporatedto dryness under a gentle stream of nitrogen. To this tube was added 30μg of rhBMP-2, sufficient MFR 00842 buffer to bring the volume up to 50μL, and 1-2 mCi of carrier-free ¹²⁵I (Dupont NEN Research Product,Boston Mass.). This solution was incubated at room temperature for 30minutes with gentile agitation. Following incubation, the solution wasadded to a NAP-5 column (Sephadex G-25, Pharmacia, Uppsala, Sweden),that had been preequilibrated with 1 column volume of MFR 00842 buffer.The column contains about 100 μL of MFR 00842 when the reaction mixturewas added. The iodinated protein was eluted from the column with MFR00842 buffer and 500 μL fractions were collected. Total ¹²⁵I content ineach fraction was determined by adding 5 μL of each fraction to apolystyrene tube containing 295 μL of bovine serum albumin (BSA, 10mg/mL) and 200 μL phosphate-buffered saline (PBS). Each tube was countedin a gamma counter for total activity. Trichoroacetic acid(TCA)-precipitable radioactivity was determined as follows: 500 μL of20% TCA was added to each tube and centrifuged at approximately 700×gfor 10 minutes. Five hundred μL of the supernatant was counted, andsoluble radioactivity was determined by the equation:

([2×supernatant CPM]+total CPM)×100

Fractions that were less than 5% soluble were pooled and stored at 4° C.Each iodination yields 30 μg of rhBMP-2 in 400 μL of buffer with a CPM(counts per minute) of approximately 3.3×10⁹.

The α-BSM test implant was prepared as described above in Example 1. Theα-BSM clinical formulation exhibited an L:S ratio of 0.85; the wetformulation had an L:S ratio of 1.0.

The ACS an absorbable collagen source (Helistat® available from IntegraLife Sciences Holdings Corp. of Plainsboro, N.J.), was prepared bypipetting a sample of the ¹²⁵I-rhBMP-2 in a buffer MFR 00926) onto thecollagen source pieces. The test implants were allowed to set for about5 minutes and implanted in the subject rabbit as soon as possible afterthis time.

Assessment of the radioactivity of the site was made using gammascintigraphy (Siemans Orbitor Gamma Camera). In order to quantify thegamma camera images, a phantom must be developed. The phantom wasdesigned based on the attenuation of the activity seen from implantationof a vial containing a known quantity of ¹²⁵I labeled rhBMP-2 at theimplantation site in a rabbit cadaver. The phantom was assessed at eachtime point to account for decay of the ¹²⁵I over time. Followingsurgical closure (5 to 10 minutes after implantation), a time zero (T0)assessment of the radioactivity at the site was made. Thereafter,animals were anesthetized, if necessary, and assessments were made.

FIG. 5 is a graph illustrating the retention of rhBMP-2 in α-BSMprepared as and measured as described above. It can be seen from thegraphs that the osteogenic composition is released from the BSM carrierat a much slower rate than from the ACS collagen sponge carrier.

FIG. 6 is a graph illustrating the release rate kinetics of rhBMP-2 fromα-BSM prepared as described above. These results indicate that theretention rate of rhBMP-2 in the α-BSM was much greater than theretention of the same rhBMP-2 in an absorbable collagen source,Helistat®.

While the embodiments of invention have been illustrated and describedin detail in the drawings and foregoing description, the same is to beconsidered as illustrative and not restrictive in character, it beingunderstood that all changes and modifications that come within thespirit of the invention are desired to be protected. In addition, allreferences cited herein are hereby incorporated by reference in theirentirety.

1-10. (canceled)
 11. An osteoinductive composition for generating a bonemass in a patient, the osteoinductive composition comprising bonemorphogenetic protein 2 within a biodegradable carrier comprisingcalcium phosphate the biodegradable carrier being substantiallyimpermeable to efflux of said bone morphogenetic protein 2 so that theformed bone mass is confined to a volume of the carrier, wherein thebone morphogenetic protein 2 is present in an amount of 0.5 mg/ml to 4mg/ml of the calcium phosphate and the bone morphogenetic protein 2 isreleased from the biodegradable carrier as the biodegradable carrierdegrades.
 12. An osteoinductive composition of claim 11, wherein saidcomposition further comprises collagen.
 13. An osteoinductivecomposition of claim 11, wherein said calcium phosphate comprisestricalcium phosphate.
 14. An osteoinductive composition of claim 13,wherein said composition further comprises hydroxyapatite.
 15. Anosteoinductive composition of claim 11, wherein said bone morphogeneticprotein 2 is recombinant human bone morphogenetic protein
 2. 16. Anosteoinductive composition claim 11, wherein said biodegradable carrierhas a closed pore structure.
 17. An osteoinductive composition claim 11,wherein said calcium phosphate is a cement and comprises tricalciumphosphate or hydroxyapatite or combinations thereof.
 18. Anosteoinductive composition of claim 11, wherein said osteoinductivecomposition further comprises an osteoinductive factor selected from thegroup consisting of LIM mineralization protein, transforming growthfactors, insulin-like growth factors, epidermal growth factors,platelet-derived growth factors and fibroblast growth factors. 19-20.(canceled)
 21. An osteoinductive composition of claim 11, wherein saidcomposition is inserted into a disc space.
 22. An osteoinductivecomposition of claim 11, wherein said composition has a chamber forreceipt of said bone morphogenetic protein
 2. 23. (canceled)
 24. Anosteoinductive composition of claim 11, wherein less than about 50% byweight of the bone morphogenetic protein 2 is released from the carrierafter about 2 days post implantation.
 25. An osteoinductive compositionof claim 11, wherein less than about 50% by weight of the bonemorphogenetic protein 2 is released from the carrier after about 7 dayspost implantation. 26-37. (canceled)